The broad aims of the program are to analyze genetic and epigenetic mechanisms underlying normal mammalian differentiation; and to examine the developmental basis of malignancy and of genetic diseases, particularly those disorders due to specific biochemical lesions, as in certain human heritable syndromes. These goals are being pursued chiefly through experimental exploitation of the developmental capacities of mouse teratocarcinoma cells. We developmentally totipotent teratocarcinoma stem cells are injected into genetically marked early embryos at the blastocyst stage, the malignant cells become stably "normalized" and participate in embryogenesis. They can contribute not only to somatic tissue differentiation but also sometimes to germ cell formation, thus giving rise to progeny. Therefore, teratocarcinoma stem cells may be utilized as vehicles for introducing into mice specific predetermined mutations. This is accomplished by first mutagenizing the cells during in vitro culture and then subjecting them to appropriate selection or screening procedures capable of yielding clones with the mutation of interest; the mutant cells are next injected into blastocysts. By thus joining in vitro and in vivo methodologies, mouse models of human genetic diseases may be produced. Other mutations, of probable interest for analyses of developmental interactions of cells (e.g., at the cell surface) may also be introduced. The mutant genes may be either nuclear or cytoplasmic (i.e., mitochondrial), murine on foreign. Cytoplasmic markers in mitochondrial DNA will enable maternal inheritance to be analyzed in vivo, for the first time in higher organisms. Introduction and integration of genes from non-murine species will provide a means of studying gene control mechanisms in relation to tissue-specific gene expression.